Respiratory syncytial virus-associated pneumonia in primary care in Malawi

Abstract Objective To identify the prevalence of respiratory syncytial virus (RSV) in a cohort of children under 5 years of age with World Health Organization (WHO)-defined pneumonia and the factors associated with developing severe RSV-associated community-acquired pneumonia (CAP) in primary care in a single centre in Northern Malawi. Methods The BIOmarkers TO diagnose PnEumonia (BIOTOPE) study was a prospective cohort study conducted from March to June 2016 that took place in a primary care centre in Northern Malawi. Data from this study was used to identify the characteristics of children under 5 years of age who presented with RSV and WHO-defined CAP. Means, standard deviations, medians and ranges were calculated for continuous variables. A univariate logistic regression was performed to examine the potential predictor variables. Results Four hundred and ninety-four infants presented with CAP and were eligible for inclusion in the study; RSV infection was detected in 205 (41.6%) of the infants. Eight factors were associated with increased risk for RSV CAP in the univariate model: age, born at term, presenting for care in June, crowded living environment, not being exclusively breastfed, not having received zinc or vitamin A supplementation in the last six months. Infants with RSV were more likely to have an oxygen saturation ≤92% compared to infants with other causes of pneumonia and more likely to have severe pneumonia as defined by the WHO. Conclusion This study supports that RSV-associated CAP is linked to modifiable and non-modifiable risk factors; further research is indicated to determine which interventions would be most impactful. Developing and implementing an infant or maternal vaccine could be a cost-effective way to prevent RSV-associated CAP and mortality in developing nations. More research is needed to understand seasonal patterns of CAP and research over extended periods can offer valuable insights on host, environmental and pathogen-specific factors that contribute to RSV-associated CAP.


INTRODUCTION
Community-acquired pneumonia (CAP) causes substantial morbidity and mortality in children under five years of age globally, with a disproportionately large number of cases in sub-Saharan Africa [1].Respiratory viruses, particularly respiratory syncytial virus (RSV), significantly contribute to CAP and childhood mortality, as evidenced by the three million hospitalizations and 60 000 annual deaths worldwide [2].Children under six months assume half of these hospitalizations and deaths [2].In Malawi 11.9% of children with severe acute respiratory infection had RSV and this had an adjusted risk ratio of 1.9 for severe infection [3].The mean cost per RSV episode in Malawi is $62.26 USD per inpatient case, which is approximately onethird of the average monthly household income [4].Contemporary estimates of the microbiological causes of pneumonia and predictors of hospitalization from primary care settings would be of benefit since it is in the community where these patients are most likely to present [5].In particular, the analysis of the burden of RSV in the community would help determine the role of prevention strategies such as immunization [5].
Decades of surveillance data on RSV in high-income nations have demonstrated that RSV causes yearly seasonal outbreaks.The timing and duration of the outbreaks vary geographically and can be predicted by location from year to year.However, pathogen-specific data is scarce or non-existent in low-income nations.The surveillance data available on RSV is limited to hospitalized patients and incomplete due to differences in reporting and a lack of diagnostic resources available.The landmark Pneumonia Etiology Research for Child Health (PERCH) study provided insight into variables associated with developing severe RSV; however, the participants were solely hospitalized patients [6].Studies investigating the risk factors associated with increased RSV disease severity in primary care are lacking in Africa.An analysis of different host factors like age, gender, premature birth and nutritional status; environmental factors such as higher house population density and increased smoke exposure; and pathogen-specific factors that affect children with RSV would be important to help address the associated morbidity and mortality [5].
This aim of this paper is to determine the prevalence and risk factors for RSV in the primary care cohort in the BIOTOPE study (BIOmarkers TO diagnose PnEumonia)-a prospective cohort study that systematically enrolled children presenting with CAP to primary care in Northern Malawi [5,7].

METHODS
The study describes the characteristics of children under 5 years of age with RSV and WHO-defined pneumonia from the BIOmarkers TO diagnose PnEumonia (BIOTOPE) study [5].In brief, participants were patients that presented to the outpatient department of Mzuzu Central Hospital and Mapale Health Centre (a primary care centre) with WHO-defined pneumonia from March to June 2016.Both sites serve as primary care facilities for the urban area of Mzuzu in Northern Malawi.Further details on the site and methods are available in an online Supplementary file (https:// bmjopen.bmj.com/content/11/7/e046633#DC1

Statistical analysis
The overall aim of this study was to determine the prevalence and risk factors that contribute to children presenting with RSV-associated CAP.Categorical variables were expressed as percentages.Means, standard deviations, medians and ranges were calculated for continuous variables.A univariate logistic regression was performed to examine the potential predictor variables.Low birthweight included participants <2.5 kg, household crowding consisted of ≥3 people sleeping in the same room as the participant, and secondary school complete was defined as 12 years of education completed.Exclusive breastfeeding was indicated as infants exclusively breastfed until six months of age.A p-value of ≤0.05 was considered statistically significant.The data was fitted to a stepwise logistic regression model to determine variables significantly associated with a positive RSV finding.Odds ratios and 95% confidence intervals were calculated.All analyses were performed using Microsoft Excel for Mac version 16.84 (Microsoft Corporation, Microsoft Excel for Mac version 16.84, Seattle, WA, USA).

RESULTS
The study group included 494 infants with CAP, of which RSV infection was detected in 205 (41.6%) children.There were 86 (42.0%) infants who were under 12 months old in the RSV-positive group compared to 71 (24.6%) in the RSV-negative group (p < 0.001).Of study participants with health passports, 374 (94%) had the pneumococcal conjugate vaccine (PCV13).The sociodemographic, environmental and clinical characteristics of the two groups are presented in Tables 1 and 2. Children positive for RSV-associated CAP were more likely to be younger, born at term, presented in June, lived in crowded spaces, had not been exclusively breastfed, had not received zinc or vitamin A supplementation in the last six months in a univariate analysis.Children with RSV were also more likely to have had an oxygen saturation ≤92% and severe pneumonia (as defined by WHO) in a univariate analysis.
Figure 1 shows the distribution of co-detected viral pathogens in children with RSV pneumonia.Bocavirus was the commonest virus detected in RSV CAP (24%).The percentage of RSV-negative CAP concurrently infected with Bocavirus was lower at 18%.Bacteria were only found in 4.9% of participants with RSV CAP.

Predictors of RSV CAP
Multivariate regression was performed to examine the predicators of RSV-associated CAP.All variables with p < 0.05 in the univariate analysis were considered for inclusion.The month of infection, age and nonexclusive breastfeeding were most associated with RSV CAP in the univariate analysis.In June, pneumonia in a child is 1.29 times more likely to be due to RSV than other viral or bacterial pathogens.The odds of RSVassociated CAP were 1% higher in a participant who was not breastfed exclusively.Nutritional supplementation (with zinc or vitamin A) also decreased the odds of RSV-associated CAP by 7.6%.Table 3 outlines notable predictors of RSV-associated CAP.

DISCUSSION
This study examined the prevalence and predictors of RSV-associated CAP among children presenting to primary care in Malawi.RSV was one of the most commonly detected viruses in children, which matched previous studies from low-income countries [8,9].The variables most associated with contracting RSV were found to be multifactorial.Demographic factors include age 2-12 months and born at term in a univariate analysis.Age greater than 36 months was associated with reduced probability of having RSV-associated CAP.Environmental factors include month of infection and more than three people sleeping in the same room.Other factors include lack of exclusive breastfeeding, no zinc or vitamin A supplementation in the last six months.Children with RSV pneumonia were more likely, than children with pneumonia due to other pathogens to have an oxygen saturation ≤92% and severe pneumonia as defined by the WHO [7].The final regression model showed that age and month of infection were significant predictors of RSV-positive CAP when controlling for covariates.
Household crowding has been consistently associated with an increased risk of acquiring RSV-associated infection regardless of the definition of household crowding [10].RSV spreads through viral shedding under close conditions; those in close contact with RSVinfected individuals are at risk of exposure to the virus  during maximal shedding timing (early in infection) and receive higher amounts of the inoculate [10].Educational interventions which target infection prevention control measures (mask-wearing, hand washing) could decrease the risk of RSV infections in overcrowded households.
Non-breastfeeding practices have been similarly cited in the literature as posing significant risk for RSVassociated lower respiratory tract infection (LRTIs) [11,12].Mineva, et al. [11] identified that exclusive breastfeeding for >4-6 months significantly lowered hospitalization, length of stay, supplemental oxygen demand and admission to intensive care units.Education on breastfeeding principles in developing countries could serve as effective primary prevention strategies.Factors contributing to exclusive breastfeeding include the mother's age, ethnicity of the mother, sex of the infant, number of children of the mother and decisionmakers within the family.Previous studies have suggested targeted interventions to promote exclusive breastfeeding that includes community breastfeeding groups, health education and awareness campaigns, the use of mass media, and the re-examination of the Baby-Friendly Hospital Initiative and Baby-Friendly Community Initiative [13,14].) There is an increased focus on RSV given its impact on child health and health services.Until recently the only medical prevention strategy available was palivizumab, a monoclonal antibody indicated in a specific subset of infants with comorbidities or delivered preterm and more recently nirsevimab is a monoclonal antibody to the RSV fusion protein with an extended half-life that could provide promising outcomes [15].

Prevention of RSV in infants
There is an increased focus on RSV given its impact on child health and health services.Palivizumab (Synagis TM) is the current standard of preventative care licenced for infants at the highest risk for severe RSV disease.Although it is expensive and intensive to administer which is a challenge for low-resource countries, it decreases hospitalization due to RSV [23].This includes those born preterm (GA < 35 weeks), younger than six months at the start of the RSV season, and children younger than two years of age with chronic lung disease of prematurity or hemodynamically significant congenital heart disease [24].Prophylaxis is started at the beginning of the RSV season and given via intramuscular injection monthly throughout the season [25].The approximate cost of one milligram of palivizumab was 8.25 Euro, equivalent to approximately 15 113 Malawian Kwacha, in 2020 [26].A one-time vaccine may be a less costly and burdensome alternative, especially in developing nations where access to primary care is limited.
More recently, many developed nations worldwide including the European Union, the UK, Canada and the USA have authorized the use of Nirsevimab (Beyfortus TM), a long-acting monoclonal antibody to the RSV fusion protein that offers 5 months of protection for RSV in infants [24].A single intramuscular injection before a late-term or term infant's first RSV season demonstrated efficacy in protecting against medically attended RSV infections.As a result, it is recommended that all infants less than 8 months of age during their initial RSV season receive a single dose of nirsevimab [27].Additionally, children with chronic lung disease of prematurity requiring medical support in the 6 months before their second RSV season, those with severe immunocompromise, those with congenital heart disease, or children with cystic fibrosis are advised to receive nirsevimab upon entering their second RSV season [24].These recommendations could change and expand as more research specifically targets lowand middle-income countries.Nirsevimab necessitates only one payment and one visit to a primary care centre in a single season.Its affordability in low-and middle-income countries remains uncertain [24,28].Syncytial virus-associated pneumonia � 5

Maternal vaccination
Worldwide, phase 3 trials, have shown preliminary evidence of safety and efficacy of preventing medically attended severe RSV-associated LRTI in infants following maternal vaccination with a bivalent RSV prefusion F protein-based (RSVpreF) vaccine at 24-36 weeks of pregnancy [28,29].Immunizing mothers with RSVpreF vaccines was found to produce neutralizing antibody responses approximately seven weeks after immunization and could adequately transfer transplacentally [29].
The MATISSE study [28] on RSVpreF vaccine involved patients from Gambia and South Africa (15.7% of total study population).
The evidence is promising thus far, but future studies must address vaccine efficacy in the long term and the logistical approach for its use in sub-Saharan Africa primary care settings.As the research on vaccinating healthy pregnant mothers continues, so should consideration about implementing these as preventative measures in the primary care setting of sub-Saharan African countries.As can be seen in this study there is high uptake of other vaccines in the current programme which highlights the potential to introduce these vaccines into the Malawian vaccination schedule [28].
Presently, RSV vaccinations are available in specific populations within high-income countries.For adults aged 60 or older, Arexvy is administered as a twovaccine series.Additionally, pregnant women can receive Abrysvo as an RSV vaccine during weeks 32 through 36 of pregnancy [30].An RSV vaccine has not yet been approved for use in children [30,31].A recent systematic review highlighted the cost and requirement of access to a dose every month mean that palivizumab is a less appealing option in low-income and middleincome countries and highlighted that the uncertainty as to whether these interventions will be cost-effective in low-and middle-income settings at current prices.Further work has highlighted that this does depend on cost.For maternal vaccination, a dose price of 40 USD may be cost-effective for Kenya and South Africa and a dose price of 10 USD cost saving in South Africa [32,33].A table summarizing these strategies is available in the Supplementary file.
Vitamin A deficiency can lead to heightened susceptibility to infections, resulting in increased morbidity and mortality [16].In Malawi, the prevalence of vitamin A deficiency has decreased due to the introduction of vitamin A-fortified staple foods and biannual highdose vitamin A supplementation.However, this success has raised concerns about potential vitamin A excess, prompting a re-evaluation of the universal approach to vitamin A supplementation [17].Similarly, zinc is involved in multiple body processes including immune system performance and DNA replication [18].Zinc deficiency in early childhood has been linked to increased risk of diarrhoea and pneumonia [18,20].During the first six months of breastfeeding, infants receive sufficient zinc intake.However, after this period, infants in sub-Saharan Africa predominantly rely on plant-derived food sources, putting them at risk of zinc deficiency.There has been evidence to support the use of zinc supplementation in RSV and other respiratory illnesses in children with one study demonstrating a 21% reduction in pneumonia incidence in children 12-59 months of age [18,19].Previous research has demonstrated that exposure to environmental tobacco smoke puts infants and young children at an increased risk of developing severe RSV [21,22].Our study did not confirm the finding; however this is likely due to fact that only a small percentage (42/494 or 8.5%) of participants were exposed tobacco smoke; and none of the participants under 12 months of age were exposed to tobacco smoke.Thus interventions to minimize tobacco smoke exposure may not be as valuable in Northern Malawi.

Limitations
Participants were recruited for our study from March to June 2016.The study was conducted during one season in an urban area with high immunization rates believed to be representative of Malawi and the Mzuzu region [5]; for this reason, we believe our study's findings may be generalized and applied to Malawi as a nation.The results of this study may also be cautiously generalizable to other countries of sub-Saharan Africa.It has been noted that significant variability in infection rates exists depending on the season; had this study been conducted throughout a different season or for 12 months, the results may have differed [5].RSV season in Southern Africa has been cited as lasting from October to May, peaking in March [18].More research is required to adequately compare RSV's seasonal patterns and other pneumonia causes.Our study found that RSV was most prevalent during June, while other causes of pneumonia were evenly distributed across the four-month study period.It is worth noting, however, that the recent COVID-19 pandemic and its associated restrictions may change the apparent seasonal patterns of RSV [19].In particular, delayed RSV outbreaks have been observed due a lack of exposure from the previous season resulting in decreased protective immunity within communities [19].

Figure 1 .
Figure 1.Distribution of other viral pathogens in children with RSV-associated pneumonia, compared to the total population studied.

Table 2 .
Clinical features and severity a RSV -Respiratory Syncytial Virus; CAP -Community acquired pneumonia; OR -Odds ratio